Rapid recruitment of BRCA1 to DNA double-strand breaks is dependent on its association with Ku80.
BRCA1 is the first susceptibility gene to be linked to breast and ovarian cancers. Although mounting evidence has indicated that BRCA1 participates in DNA double-strand break (DSB) repair pathways, its precise mechanism is still unclear. Here, we analyzed the in situ response of BRCA1 at DSBs produced by laser microirradiation. The amino (N)- and carboxyl (C)-terminal fragments of BRCA1 accumulated independently at DSBs with distinct kinetics. The N-terminal BRCA1 fragment accumulated immediately after laser irradiation at DSBs and dissociated rapidly. In contrast, the C-terminal fragment of BRCA1 accumulated more slowly at DSBs but remained at the sites. Interestingly, rapid accumulation of the BRCA1 N terminus, but not the C terminus, at DSBs depended on Ku80, which functions in the nonhomologous end-joining (NHEJ) pathway, independently of BARD1, which binds to the N terminus of BRCA1. Two small regions in the N terminus of BRCA1 independently accumulated at DSBs and interacted with Ku80. Missense mutations found within the N terminus of BRCA1 in cancers significantly changed the kinetics of its accumulation at DSBs. A P142H mutant failed to associate with Ku80 and restore resistance to irradiation in BRCA1-deficient cells. These might provide a molecular basis of the involvement of BRCA1 in the NHEJ pathway of the DSB repair process.
Wei
Leizhen
L
Department of Clinical Oncology, Tohoku University, 4-1 Seiryomachi Aoba-ku, Sendai 980-8575, Japan.
Lan
Li
L
Hong
Zehui
Z
Yasui
Akira
A
Ishioka
Chikashi
C
Chiba
Natsuko
N
D034961:Antigens, Nuclear / Q000235:genetics / Q000378:metabolism*; D019313:BRCA1 Protein / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004247:DNA / Q000528:radiation effects*; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005260:Female; D006657:Histones / Q000235:genetics / Q000378:metabolism; D006801:Humans; D000072200:Ku Autoantigen; D007834:Lasers; D020125:Mutation, Missense; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism
D034961:Antigens, Nuclear; D019313:BRCA1 Protein; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D006657:Histones; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D025521:Tumor Suppressor Proteins; D004247:DNA; C102449:BARD1 protein, human; D044767:Ubiquitin-Protein Ligases; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen